Two-photon excited hemoglobin fluorescence provides contrast mechanism for label-free imaging of microvasculature in vivo

Direct visualization of microvasculature provides significant insights in microcirculation and critically impacts the diagnosis and treatment of microcirculatory diseases. Recently, we discovered that the high-energy Soret fluorescence of hemoglobin peaked at 438?nm with an extremely short lifetime becomes strongly visible under two-photon excitation. Based on the distinct spectral and temporal characteristics of hemoglobin fluorescence, we demonstrated that two-photon fluorescence microscopy could become a powerful tool for label-free in vivo imaging of microvasculature in tissue.     

采用氧合血红蛋白吸收带的漫反射光谱比率R（540）/R（575）检测浅表性膀胱癌

采用氧合血红蛋白在540和575 nm吸收带的漫反射光谱比率(R540/R575)对人浅表性膀胱癌进行了鉴别诊断,实验采用带积分球附件的分光光度计获取组织的漫反射光谱。结果表明:正常膀胱的粘膜/粘膜下层组织的漫反射光谱比率(R540/R575)在组织离体后2 h的平均值为111%,3 h的平均值为107%,4 h的平均值为104%,5 h的平均值为102%,而浅表性的膀胱癌的粘膜/粘膜下层组织的漫反射光谱比率(R540/R575)在组织离体后2 h的平均值为98.4%,3 h的平均值为95.5%,4 h的平均值为93.1%,5 h的平均值为91.6%,正常膀胱与浅表性膀胱癌的粘膜/粘膜下层组织在组织离体后2,3,4和5 h的漫反射光谱比率(R540/R575)均有显著性的差异(p<0.05),分别为12.6%,11.5%,10.9%和10.4%。这结论为快速、低成本、非入侵的浅表性膀胱癌的光活检提供了一种新的方法。     

Spectroscopic analysis of bladder cancer tissues using Fourier transform infrared spectroscopy

Bladder cancer is one of the most common cancers in Africa. It takes several days to reach a diagnosis using histological examinations of specimens obtained by endoscope, which increases the medical expense. Recently, spectroscopic analysis of bladder cancer tissues has received considerable attention as a diagnosis technique due to its sensitivity to biochemical variations in the samples. This study investigated the use of Fourier transform infrared (FTIR) spectroscopy to analyze a number of bladder cancer tissues. Twenty-two samples were collected from 11 patients diagnosed with bladder cancer from different hospitals without any pretreatment. From each patient two samples were collected, one normal and another cancerous. FTIR spectrometer was used to differentiate between normal and cancerous bladder tissues via changes in spectra of these samples. The investigations detected obvious changes in the bands of proteins (1650, 1550 cm^–1), lipids (2925, 2850 cm^–1), and nucleic acid (1080, 1236 cm^–1). The results show that FTIR spectroscopy is promising as a rapid, accurate, nondestructive, and easy to use alternative method for identification and diagnosis of bladder cancer tissues.     

人膀胱粘膜/粘膜下层组织病变致组织漫反射光谱的变化

研究了人正常膀胱和浅表性膀胱癌的粘膜/粘膜下层组织在300～900nm光谱范围的漫反射光谱特性及其差异,实验采用带积分球附件的分光光度计获取组织的漫反射光谱.结果表明,在300～900 nm,正常膀胱的粘膜/粘膜下层对任一个波长的漫反射率都明显地较癌变的粘膜/粘膜下层对相应波长的要大.正常膀胱的粘膜/粘膜下层的漫反射光谱的峰分别在370 nm、520 nm、550 nm和660nm,其峰值分别为52.4%、60.7%、56.1%和75.5%.而癌变的粘膜/粘膜下层的漫反射光谱的峰分别在320 nm、520 nm、550 nm和660 nm,其峰值分别为43.7%、33.4%、30.6%和70.2%.正常膀胱的粘膜/粘膜下层的漫反射光谱在370 nm处有一个峰,而癌变的粘膜/粘膜下层的漫反射光谱在370 nm处没有峰,320 nm处有一个峰.而正常膀胱的粘膜/粘膜下层的漫反射光谱在320 nm处没有峰.膀胱的粘膜/粘膜下层病变导致组织的漫反射光谱在520 nm、550 nm和660 nm处的峰的峰值分别减小了45.0%、45.5%和7.02%.说明膀胱的粘膜/粘膜下层病变导致组织的组分和结构尤其足组织中的氧合血红蛋白和去氧血红蛋白的含量发生了明显的改变.     

Nonionizing photoacoustic cystography in vivo

We demonstrate the feasibility of a novel and nonionizing process for bladder imaging in vivo, called photoacoustic cystography (PAC). Using a photoacoustic imaging system, we have successfully imaged a rat bladder filled with clinically used Methylene Blue (MB) dye. An image contrast of ~8 was achieved. Further, spectroscopic PAC confirmed the accumulation of MB in the bladder. Using a laser pulse energy of less than 1 mJ/cm2 (1/20 of the ANSI safety limit), a deeply (1.2 cm) positioned bladder in biological tissues was clearly visible in the PA image. Our results suggest that PAC can potentially provide a nonionizing, relatively cheap, and portable tool for bladder mapping. Among our clinical interests, nonionizing PAC with an injection of MB can potentially monitor vesicoureteral reflux in children.     

肿瘤和正常组织的FT-Raman与FT-IR研究

本文运用ＦＴ－Ｒａｍａｎ光谱法研究肠、胃、十二指肠、口腔腮腺等部位正常组织与肿瘤组织，发现正常组织与肿瘤组织在Ｒａｍａｎ光谱上表现出较为明显的差别，这一结果与我们先前所得到的红外光谱结果相一致。此结果说明Ｒａｍａｎ光谱亦有潜力发展成为一种活体、原位、无损诊断肿瘤的新方法。     

Enhancing early bladder cancer detection with fluorescence-guided endoscopic optical coherence tomography

We report an experimental study of the possibility of enhancing early bladder cancer diagnosis with fluorescence-image-guided endoscopic optical coherence tomography (OCT). After the intravesical instillation of a 10% solution of 5-aminolevulinic acid, simultaneous fluorescence imaging (excitation of 380-420 nm, emission of 620-700 nm) and OCT are performed on rat bladders to identify the photochemical and morphological changes associated with uroepithelial tumorigenesis. The preliminary results of our ex vivo study reveal that both fluorescence and OCT can identify early uroepithelial cancers, and OCT can detect precancerous lesions (e.g., hyperplasia) that fluorescence may miss. This suggests that a cystoscope combining 5-aminolevulinic acid fluorescence and OCT imaging has the potential to enhance the efficiency and sensitivity of early bladder cancer diagnosis.     

Visualization of microvasculature and thrombi by X‐ray phase‐contrast computed tomography in hepatocellular carcinoma

Visualization of the microvascular network and thrombi in the microvasculature is a key step to evaluating the development of tumor growth and metastasis, and influences treatment selection. X‐ray phase‐contrast computed tomography (PCCT) is a new imaging technique that can detect minute changes of density and reveal soft tissues discrimination at micrometer‐scale resolution. In this study, six human resected hepatocellular carcinoma (HCC) tissues were investigated with PCCT. A histological stain was added to estimate the accuracy of PCCT. The results showed that the fine structures of the microvasculature (measuring 30–100 µm) and thrombi in tiny blood vessels were displayed clearly on imaging the HCC tissues by PCCT. Moreover, density distributions of the thrombi were obtained, which could be reliably used to distinguish malignant from benign thrombi in HCC. In conclusion, PCCT can clearly show the three‐dimensional subtle structures of HCC that cannot be detected by conventional absorption‐based computed tomography and provides a new method for the imageology of HCC.     

Diagnosis of colon cancer using frequency domain fluorescence imaging technique

Early detection and treatment of colon cancer has been associated with better disease prognosis. Conventional and reported optical techniques have limitations in detecting early stages of colon cancer growth. In this paper, a homodyne signal processing assisted frequency domain (FD) fluorescence imaging methodology is proposed for the early diagnosis of colon cancer. Simulated phantom tissues representing the biopsy samples at different stages of colon cancer growth are prepared and used for the imaging study. Selective imaging of healthy and diseased sites simulated in the samples was achieved even for fluorescence emissions having close lifetimes and wavelength values. Possible extension of the methodology for in vivo investigations is also discussed.     

膀胱肿瘤离体组织的拉曼光谱学研究

使用激光共聚焦显微拉曼光谱仪测取膀胱肿瘤和正常膀胱组织的拉曼特征谱,应用主成分分析/支持向量机(principal component analysis,PCA/support vector machines,SVM)分类器对数据进行判别分析,最后使用弃一交叉验证法(leave-one-out cross validation,LOOCV)测试判别结果的准确度。结果发现膀胱肿瘤组织与正常膀胱组织的拉曼光谱存在明显差异,肿瘤组织在782和1 583cm-1等核酸特征谱带处峰高显著增强,而正常组织在1 061,1 295,2 849,2 881cm-1等蛋白质和脂质特征谱带处峰高显著增强。PCA/SVM可良好区分膀胱肿瘤组织和正常膀胱组织的拉曼光谱,LOOCV测试分类器显示肿瘤诊断的敏感度86.7%、特异度87.5%、阳性预测值92.9%、阴性预测值77.8%。由此得出结论:拉曼光谱可以良好诊断膀胱肿瘤的体外组织,展现了优越的临床应用前景。     

基于拉曼光谱和多元统计分析技术的结肠癌早期诊断

为了寻找结肠癌的病理发展规律在拉曼光谱属性上的体现,采用共聚焦显微拉曼光谱仪对60例离体的正常结肠组织、腺瘤性息肉和腺癌组织的近红外拉曼光谱进行对比检测,初步探讨了三类组织的拉曼光谱特征及其改变的规律。结果表明:三类组织拉曼光谱的差异明显存在于830,855,1 032,1 210,1 323,1 335,1 445,1 450,1 655 cm^-1处,腺瘤性息肉的光谱大体位于正常组织与腺癌组织之间。以组织病理诊断为金标准,主成分分析结合线性判别分析技术建立的诊断算法区分3类组织的灵敏度分别为96.9%、85.7%和97.3%,特异性分别为82.8%、90%和92.3%。因此,拉曼光谱有潜力在分子水平上区分正常结肠组织、腺瘤性息肉和腺癌组织,有望成为结肠癌早期诊断的一种有效方法。     

乳腺癌的傅里叶变换红外光谱成像研究

傅里叶变换红外光谱(FTIR)成像技术可同时获得组织样本的空间分布信息和红外光谱信息。本文利用FTIR成像技术对离体原位乳腺组织样本进行原位研究,通过特征谱带吸光度比值(面积比值)图像结合FTIR光谱分析研究健康和癌变乳腺组织的差异。发现乳腺癌组织的特征带吸光度比值A1087/A1455、A1238/A1455、A1550/A1455、A1650/A1455相对于健康组织明显增大,A1160/A1087、A1740/A1550则减小。研究结果论证了乳腺组织癌变过程中大分子含量的变化规律,体现了FTIR成像技术在乳腺癌诊断方面的准确性、直观性和可行性,其有潜力成为基础研究和临床诊断的一种准确有效的技术手段。     

Optical-resolution photoacoustic microscopy for in vivo imaging of single capillaries

Capillaries, the smallest blood vessels, are the distal end of the vasculature where oxygen and nutrients are exchanged between blood and tissue. Hence, noninvasive imaging of capillaries and function in vivo has long been desired as a window to studying fundamental physiology, such as neurovascular coupling. Existing imaging modalities cannot provide the required sensitivity and spatial resolution. We present in vivo imaging of the microvasculature including single capillaries in mice using optical-resolution photoacoustic microscopy (OR-PAM) developed in our laboratory. OR-PAM provides a lateral resolution of 5 microm and an imaging depth >0.7 mm.     

癌变与正常直肠组织的核磁共振氢谱差异的研究

核磁共振(NMR)波谱法是一种表征分子结构、组成变化的有效手段,能够得到生物组织中的核酸、蛋白质、脂类和糖类等生物大分子的含量、分子的空间排列和结构特征等信息,可以从分子水平上研究肿瘤与对照组织之间的差别,更深入地揭示肿瘤的发生、发展中产生的各种分子水平变化。文章对9例直肠癌组织及癌旁正常组织进行了核磁共振氢谱的测定。结果表明,直肠正常和癌变组织的核磁共振氢谱存在显著的差异,这可以通过对谱图特征峰区域积分面积相对比值的差别看出来。直肠正常组织核磁共振氢谱中A0.9/A3.0,A1.3/A3.0,A2.0/A3.0,A1.3/A0.9及A4.1/A3.0值较相应的癌组织偏大,直肠癌组织核磁共振氢谱中A3.2/A3.0值较相应正常直肠组织偏大。通过这些差异核磁共振氢谱技术有可能发展成为早期诊断直肠癌组织的一种方法。     

Certain Trace Metal Disorders in Urinary Bladder of Bilharzial Patients suffering Bladder Carcinoma

The objective of the present work is to investigate any possible role which could have been played by the long-term treatment with antimony as an antibilharzial drug, and the development of urinary bladder carcinoma in patients suffering bilharzial complications. The present work has been planned to investigate the concentration levels of copper, zinc and antimony in urinary bladder tissues sampled from normal, bilharzial and bladder cancer patients subjected to surgical intervention. Both atomic absorption spectrophotometry and neutron activation analysis have been utilized for elemental estimations. The data obtained showed significant increase in the concentration levels of the three investigated metals in bladder cancer tumours as compared with their control levels under either normal or cancer-free bilharzial complications. The results have been statistically evaluated and discussed in view of the relevant literature. It has been concluded that the contributing role of the antibilharzial antimonial treatment in development of urinary bladder cancer cases cannot be overlooked and awaits further investigation.     

生物医学光声成像的研究进展

光声成像是21世纪初发展起来的新兴的生物医学成像技术,它同时具备光学成像和声学成像两者的优点,因而备受关注。本文对生物医学光声成像的发展进行了综述。首先,介绍了光声成像的特点以及相对于广泛应用的光学成像技术和声学成像技术的优点;其次,在成像原理上解释了光声成像优点的成因,并介绍了光声断层成像和光声显微镜这两种典型的光声成像技术;再次,详细介绍了多尺度的光声图像分辨率和成像深度,以及多信息维度的光声成像参数;最后,展望了光声成像在生物医学领域的应用潜力并讨论了其局限性。     

High-resolution in vivo imaging of blood vessels without labeling

We demonstrate that both oxyhemoglobin and deoxyhemoglobin have sequential two-color, two-photon absorption properties that can serve as endogenous contrasts in microvasculature imaging. Using a sensitive modulation transfer technique, we are able to image hemoglobin in red blood cells with micrometer resolution, both in vitro and in vivo. We show that excellent contrast from hemoglobin without any labeling can be obtained in tissue.     

High-resolution imaging characterization of bladder dynamic morphophysiology by time-lapse optical coherence tomography

We report an experimental study of the possibility of high-speed optical coherence tomography (OCT) for high-resolution imaging characterization of detrusor dynamic morphophysiology and analysis of the mechanisms that lead to geriatric incontinence (GI). The spontaneous contractility of intact fresh rabbit bladders was imaged with two-dimensional (2D) OCT ex vivo at up to 8 frames/s. The time-lapse 2D OCT images were postprocessed by image segmentation and fast-Fourier-transform analysis to characterize the dynamic morphological changes of the bladder contractility. In addition, we studied young and aging rat bladders to analyze the differences in dynamics. Preliminary results of our ex vivo study reveal that time-lapse OCT can track the contractile waves of bladders at high spatial resolution and characterize their dynamic morphophysiology in terms of amplitude, phase, and frequency. The results suggest that time-lapse OCT has the potential to act as a detrusor optical biopsy to enhance the diagnosis of detrusor dysfunction and thus of the mechanisms that lead to GI.     

光声内窥镜系统在人体直肠癌离体组织中的实验研究

研制了基于4阵列传感器的光声内窥镜探头,并基于此搭建了光声内窥镜成像系统,开展了仿体实验成像研究,通过内窥镜探头4个阵元位置激光吸收强度的变化情况来分析仿体中吸收体的位置,论证了其定位能力,利用光声内窥镜系统对人体离体的正常组织、直肠癌早期组织进行成像研究;通过对不同阵元位置的光吸收强度分布进行统计分析,证明了光声内窥镜对早期直肠癌组织和正常组织的辨别能力,此项技术有望提高直肠癌早期诊断的准确率,具有潜在的临床应用前景。     

核磁共振波谱在肿瘤诊疗中的应用研究进展

在介绍肿瘤样品代谢物的核磁共振波谱技术的研究方法的基础上,从离体组织和活体组织两个方面综述核磁共振波谱(NMR)在诊断肿瘤方面的应用进展,分析了它在肿瘤诊疗中的应用前景。在离体组织方面,人们通过1H和31P-NMR谱观测病人的体液样品、培养的细胞、切除的组织等来研究脂质、磷脂等代谢物的分布,观测肿瘤与对照组织之间的差别。其中利用组织提取物的方法能够得到分辨率较高的图谱,非常适合应用于肿瘤诊断和治疗方法的研究。高分辨魔角旋转(HR-MAS)的方法在肿瘤诊断研究方面展现出新的生命力,利用高分辨魔角旋转技术可以直接得到组织细胞中很多分子水平的代谢物结构和组成信息,因此它在癌症的早期诊断中具有很好的前景。在活体核磁共振波谱诊断肿瘤方面,主要应用1H和31P核磁共振波谱,结合MRI为非侵入性肿瘤分析提供了一种临床可用的方法。MRI与MRS技术的结合将使核磁共振波谱在医学领域有更大的应用空间。